Virion host shutoff protein (VHS), the product of UL41 gene of herpes simplex virus 1(HSV-1) is the major viral suppressor of host responses to infection. VHS is brought into the infected cells by the virion and its primary function is to degrade mRNA. Our studies on VHS have completely revolutionized our understanding perceptions of the functions of the protein. Specifically: (i) We have developed a procedure for solubilization and purification of VHS to homogeneity. Purified VHS is an endoribonuclease that cleaves synthetic RNA with the specificity of RNase A in the absence of cellular or viral proteins. (ii) Earlier reports indicated that at mid and late times after infection VHS is neutralized by the viral protein VP16. We showed that neutralization required the formation of a complex that includes VP22 in addition to VP16. (iii) Earlier reports indicated that the degradation of mRNA mediated by VHS is nonselective. We have identified and quantified the VHS- dependent degradation of 3 classes of mRNAs as follows: Class A consists of mRNAs that are normally stable (e.g. actin, GAPDH mRNAs, normal half life >12 h). In infected cells these mRNAs are degraded 5' to 3' within 30 minutes. Class B consists of stress response mRNAs (e.g. c-fos, cox-2, I:B1 and IEX-1 mRNAs) induced immediately after infection. These mRNAs contain AU-rich elements within their 3' UTRs. Typical half life of these mRNAs in mock infected cells is 30 to 45 min. Early after infection, these mRNAs are deadenylated and cleaved 5' to AU-rich elements. The residual portions linger with a half life >3 hrs. At late times (6+ h after infection) degradation of these mRNAs is dependent on ICP27. Both class A and class B mRNAs are degraded in polyribosomes. Class C includes mRNAs that are not degraded. The products of these mRNAs include tristetraprolin, a stress inducible protein that binds to AU-rich elements and sequesters the mRNAs to exosomes for 3' to 5' degradation and GADD452 a transcriptional factor. (iv) Earlier reports indicated that VHS binds 3 components of eIF4F complex, i.e. eIF4H, eIF4B and eIF4AII and VP16. Our studies have shown that VHS also interacts physically with tristetraprolin, ICP27, and through VP16 with VP22. The objective of the research proposal is to test a model of the degradation of the mRNAs in classes A and B. The model proposes that VHS degrades mRNAs in polyribosomes as reported, that it binds to a still to be defined component of eIF4F and mimics the function of the decapping enzymes by cleaving stable mRNAs downstream of the cap structure. The residual portion is then sequestered in P-bodies and rapidly degraded. In the case of class B mRNAs, VHS binds to tristetraprolin bound to AU-rich element mRNA and cleaves the mRNA 5' to the AU-rich elements. In the absence of bound tristetraprolin, the residual mRNA is not recruited promptly for degradation 3' to 5' by exosomes. We also propose to discriminate between 2 alternative potential functions of ICP27 in enabling mRNA degradation late in infection. Finally, given the multiple functions of VHS, the question arises as to the extent to which each function contributes to the pathogenic potential of HSV-1.